The human met protooncogene product (Met or Met protein) a member of the family of tyrosine kinase growth factor receptors, was identified via the activated oncogene tpr-met (C. S. Cooper et al., Nature 311: 29 (1984); M. Park et al., Cell 45: 895 (1986)). Met is synthesized as a glycosylated 170-kD precursor and cleaved in the external (ligand binding) domain to yield a mature disulfide-linked xcex1- (50-kD), xcex2- (140-kD) heterodimer (C. S. Cooper et al., Nature 311: 29 (1984); S. Girodano et al., Oncogene 4: 1383 (1989); D. L. Faletto et al., Oncogene, in press (1992)). The Met receptor is expressed in a wide variety of tissue and cell types, but the highest levels are found in epithelial cells (M. F. Di Renzo et al., Oncogene 6: 1997-2003 (1991); A. Iyer et al., Cell Growth and Diff. 1: 87 (1990)).
Hepatocyte growth factor (HGF) was first purified from human and rabbit plasma and rat platelets on the basis of its ability to stimulate mitogenesis of rat hepatocytes (E. Gohoda et al., J. Clin. Invest. 81: 414 (1988); R. Zarnegar and G. Michalopoulos, Cancer Res. 49: 3314 (1989); T. Nakamura et al. FEBS Lett. 224: 311 (1987)). Thus, HGF may act as a humoral factor promoting liver regeneration after partial hepatectomy or liver injury (G. K. Michalopoulos, FASEB J. 4: 176 (1990)). The same factor was purified from human fibroblast culture medium and shown to act on melanocytes and a variety of epithelial and endothelial cells (J. S. Rubin et al., Proc. Natl. Acad. Sci. U.S.A. 88: 415 (1990)). Together with evidence of HGF expression in several organs (J. S. Rubin et al., Proc. Natl. Acad. Sci. U.S.A. 88: 415 (1990); K. Tashiro et al. Proc. Natl. Acad. Sci. U.S.A. 87: 3200 (1990); R. Zarnegar et al., Proc. Natl. Acad. Sci. U.S.A. 87: 1252 (1990); T. Kinoshita et al. Biochem. Biophys. Res. Comm. 165: 1229 (1989)), these findings indicate that HGF may also act as a paracrine mediator of proliferation for a broad spectrum of cell types. Molecular cloning of HGF revealed a remarkable structural homology to plasminogen and related serine proteases (J. S. Rubin et al., Proc. Natl. Acad. Sci. U.S.A. 415 (1990); T. Nakamura et al., Nature 342: 440 (1989); K. Miyazawa et al., Biophys. Res. Comm. 163: 967 (1989)).
HGF is structurally related to the family of serine proteases that includes plasminogen, prothrombin, urokinase, and tissue plasminogen activator (J. S. Rubin et al., Proc. Natl. Acad. Sci. U.S.A. 88: 415 (1990)); T. Nakamura et al., Nature 342: 440 (1989)). As defined in the present invention, HGF includes a variant of HGF previously characterized as a broad-spectrum mitogen called plasminogen like growth factor (PLGF). Several proteases, including members of the serine protease family, stimulate DNA synthesis presumably through a proteolytic mechanism similar to tryptic activation of the insulin receptor (S. E. Shoelson et al. J. Biol. Chem. 263: 4852 (1988)). Only urokinase has been found to associate with a specific cell-surface receptor, which itself bears no homology to any known tyrosine kinase receptors (A. L. Roldan et al., EMBO J. 9: 467 (1990)).
U.S. patent application Ser. No. 07/642,971, incorporated by reference above, describes the complex comprising HGF and met protooncogene protein and identifies the met protooncogene as the receptor for HGF.
Scatter factor (SF) originally had been considered to be related to but different from HGF, SF being associated with cell motogenicity (motility), and HGF being associated with cell mitogenicity (growth). However, recently it has been shown that HGF is identical to SF, and this factor is now referred to as xe2x80x9cHGF/SFxe2x80x9d (E. Gerardi et al., Cancer Cells 3: 227 (1991); E. M. Rosen et al., Cell Growth and Diff. 2: 603 (1991); L. Naldini et al., EMBO J. 10: 2876 (1991b); K. M. Weidner et al., Proc. Natl. Acad. Sci. USA 88: 7001 (1991)) and has been independently shown both to promote epithelial cell motility (scattering) and to cause certain epithelial cell lines to become invasive in in vitro assays (E. M. Rosen et al., Cell Growth and Diff. 2: 603 (1991); M. Stoker et al., Nature 327: 239 (1987)).
The scattering response of HGF/SF establishes that this factor is responsible for cell motility and differentiation (G. F. Vandewoude Japan. J. Can. Res. 83: cover (1992)). For instance, MDCK cells grown in collagen gels in the presence of HGF/SF form bracing tubules, suggesting that the three-dimensional geometry of cell-substrate interactions directs MDCK cells to organize into tubules in response to HGF/SF (R. Montesano et al., Cell 67: 901 (1991)).
It is known that mammary gland epithelia undergo developmental changes during pregnancy to become secretory. In culture, mammary epithelial cells can regain their differentiated phenotype only when appropriate hormonal and substratum conditions are provided (M. J. Bissell et al., The Mammary Gland, M. Neville and C. Daniel, Eds. (Plenum Press Publishing Corp., New York, pp. 97-146 (1987); C. H. Streuli et al., J. Cell Biol. 115: 1383 (1991)). Thus, a need exists to determine whether there is met expression in normal breast duct cells and to determine the role that HGF/SF and met play in the differentiation of ductal epithelium in the mammary gland. In this connection, a need also exists to determine whether there is a general phenotype in epithelial cancers arising from organs that normally involve met expression for differentiation.
The work of Bieche et al., The Lancet, 339: 139 (1992) has shown that the loss of heterozygosity on chromosome 7q is associated with aggressive primary breast cancer. Specifically, Bieche et al. used the c-met proto-oncogene probe, which detects sequences on chromosome 7q31, to analyse tumor and blood leucocyte DNA samples from 245 patients with primary breast cancers. The pmet H polymorphic probe detected a high frequency of allele loss (40-50%) among the 121 informative (heterozygous) patients. This genetic alteration was not significantly associated with standard prognostic features including tumor size, histopathologic grade, and lymph-node or steroid receptor status. However, patients with loss of heterozygosity on chromosome 7q31 in primary tumor DNA had significantly shorter metastasis-free survival and overall survival after surgery than patients without this alteration. Based upon this observation, Bieche et al. hypothesized that the region on chromosome 7 detected by the probe might be the site of a breast cancer or metastasis suppressor gene. Bieche et al. did not identify a specific genetic locus responsible for poor prognosis in breast cancer patients.
Thus, one object of the present invention is to provide a method for predicting the progression of breast cancer based upon the presence of Met, met DNA and met mRNA in normal and breast cancer tissue. Such a method would be advantageous in the management of breast cancer therapy, used either alone or in conjunction with other prognostic features such as tumor size, histo- and cytopathological grade and lymph node or steroid receptor status.
The present invention relates to a method for predicting breast tumor progression by determining one or more of met DNA abundance, met mRNA abundance, or Met protein abundance in normal breast tissue, wherein a higher abundance of met genes, met mRNA, or Met protein in the normal tissue than in the tumor tissue indicates a high likelihood of tumor metastisis.